Apolipoprotein (apo) E4 is an established risk factor for neurodegenerative disease, including Alzheimer's disease (AD), and for poor outcome from head trauma and stroke. However, the mechanism underlying this increased risk remains elusive. Since protein function is directly related to protein structure, we have focused on determining the structural features that distinguish the apoE isoforms to gain insight into how these differences relate to the mechanism for the different effects of the isoforms in neurodegeneration. Our structural and mutagenesis studies established that apoE contains two structural domains. In apoE4, but not apoE3 and apoE2, the two domains interact. Our working hypothesis is that this unique structural property of apoE4 has a major influence on its functional properties, including lipid transport, metabolism, and mechanisms by which apoE4 contributes to neurodegeneration and heart disease. The aims in this application are designed to test this hypothesis in the context of neurodegeneration and AD using in vitro model systems and a novel apoE mouse model, in which domain interaction was engineered into mouse apoE in by gene targeting (Arg-6 1 mouse apoE). In Specific Aim 1, we will test the hypothesis that domain interaction determines the lipid-binding properties of Arg-6 1 mouse apoE and human apoE4. In Specific Aim 2, we will test the hypothesis that domain interaction in Arg-61 mouse apoE influences the type and composition of lipoprotein particles secreted by cultured primary astrocytes. In Specific Aim 3, we will test the hypothesis that domain interaction in Arg-6 1 mouse apoE decreases neuronal outgrowth in cell and organ culture systems. In Specific Aim 4, we will test the hypothesis that domain interaction in Arg-6 1 mouse apoE4 contributes to neurodegeneration. The results from these studies have the potential to provide clues into the mechanism by which apoE4 contributes to neurodegeneration and to identify therapeutic targets based on isoform-specific effects.